Modeling and experimental approaches for investigating lipid bilayer heterogeneity

Kevin Bradley Towles

doi:10.17918/etd-1798

The idea of lateral heterogeneity in native cell membranes is not a new one; however, the research in this field over the last several decades has uncovered the complexity of lipid and protein interactions which govern many cellular processes. Research in this area is increasingly motivated by the concept of the "lipid raft," membrane domains enriched in cholesterol and sphingomyelin, as a functional component of the membrane. Their involvement in processes including signal transduction, molecular tracking, membrane fussion/sion, signaling, and others has been proposed through exhaustive studies. While the existence and importance of lipid rafts is now generally accepted, their characterization is exceedingly difficult. Many experimental methods point to the size of rafts being on the order of tens of nanometers, but there is a lack of experimental and theoretical understanding of these structures at such small length scales. With recent evidence suggesting the exquisite sensitivity of these systems to perturbation by "impurities," the need for non-perturbing methods of characterization and fundamental models describing the membrane thermodynamics is becoming more pronounced. This work presents a new method of characterization of small membrane domains (about 50 nm or less) based on the theory of fluorescence resonance energy transfer (RET). Comparing model predictions with simulated Monte Carlo data, this work demonstrates that time-resolved RET measurements can provide detailed size information of nanoscale membrane domains with error of about 50% or less. Moreover, these measurements use fluorophore-labeled lipids in low concentration (typically much less that 1 mol %) that preferentially partition between coexisting lipid phases, which should minimally perturb the membrane. Future experiments to confirm this model and its potential are suggested. A membrane model is also developed to describe fluid-fluid heterogeneity as a function of the lipid properties in each phase. This model provides useful insight about small membrane domains by suggesting critical domain dimensions, below which domains should be highly unstable entities. The model also shows that previous estimates of line tension in model membranes may be a lower limit and that higher line tensions and the balance between the properties of each phase could drastically alter the kinetics of domain growth. Improvements to the model to include domain budding are also suggested.

Modeling and experimental approaches for investigating lipid bilayer heterogeneity

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Modeling and experimental approaches for investigating lipid bilayer heterogeneity

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